Tremendous advances have recently been made in the field of bioelectronics. Biosensors, biological microelectromechanical systems (bio-MEMs) and microfluidic devices are revolutionizing biological and medical research. Similarly, the development of biomolecular arrays and regenerative therapies are enabling unprecedented studies in fundamental biology and applications in tissue engineering, respectively.
The ability to define multiple biomolecules on a single surface while retaining their functionality is integral to the development of, for example, bio-devices, for biomolecule microarrays and in tissue engineering. Consequently, multicomponent patterning of biomolecules has become an active area of research. Patterning techniques for single component patterning have been extensively developed. However, multicomponent patterning introduces unique challenges, which have yet to be satisfactorily addressed. Multicomponent protein patterning requires the ability to define multiple biomolecules on a single substrate while avoiding nonspecific binding and retaining biomolecule integrity. All of these issues must be addressed in evaluating a patterning method.
Techniques using photolithography, soft-lithography, dip-pen lithography, and spot-arraying have all been explored and represent the principal advances in the field of multicomponent protein patterning. Photolithography is a mature patterning technique that is promising for biomolecule applications as it allows for high-resolution and precise alignment, and is a high-throughput process. The greatest disadvantage of photolithography is that it traditionally requires harsh photoresists and developers as well as heating, which often leads to denaturation of delicate biomolecules. Many clever strategies have been developed to circumvent these requirements. However, these strategies are necessarily specialized and lack universal applicability.
Soft-lithography is an inexpensive and parallel patterning process, which also has the ability to produce three-dimensional structures. However, due to the elastomeric nature of the stamps, mechanical deformation necessarily occurs and results in pattern deformation. Furthermore, soft-lithography is limited by lack of registration capabilities. Dip-pen lithography offers the ability to precisely place nanoscale arrays of proteins; though, by nature, this technique proves to be prohibitively time-consuming. Spot-arraying techniques, including ink-jet printing, are able to generate arrays with spot sizes close to 100 μm. Although this can be a very cost-effective method, resolution is too low to be practical for many applications.
Conventional lithographic patterning methods are attractive for this application, but the fact that they require the use of organic solvents and imaging materials, which are in general damaging to fragile biomolecules, is considered a technical bottleneck. Preserving the integrity of proteins during the patterning process and non-specific binding remain significant challenges to be overcome. In particular, the issue of binding multiple proteins to a single surface has yet to be satisfactorily addressed, despite substantial research on the subject. Based on the foregoing, there exists an ongoing and unmet need for a method of patterning biomaterials, and in particular making multiple overlaid patterns of biomaterials.